PubMed Health⌕ Search

Biomedical subjects

K R Porter

Publications and source records attributed to K R Porter.

At least 73 records · Page 4Linked to original sources
PubMed1982

The control of pigment migration in isolated erythrophores of Holocentrus ascensionis (Osbeck). II. The role of calcium.

The integumental pigment cells (erythrophores) of the squirrel fish, Holocentrus ascensionis, are specialized for rapid radial transport of the pigment granules contained within their cytoplasm. Pigment granules in isolated denervated erythrophores alternate spontaneously between a centrally aggregated state and a radially dispersed state. In the absence of external calcium, pigment aggregation does not occur spontaneously and cannot be induced by the aggregating agents epinephrine or high concentration of external K+. Pigment aggregation is also impaired in the presence of D600 or papaverine, compounds reported to antagonize calcium influx into the cell. Pigment aggregation can be induced by experimental elevation of the concentration of cytoplasmic free Ca2+, with a Ca-EGTA buffer system in conjunction with ionophore A23187. The threshold concentration of Ca2+ required to produce this effect is 5 X 10(-6) M. These results suggest that cytoplasmic free Ca2+ is involved in mediating pigment aggregation and that some, if not all, the Ca2+ is supplied by influx from the extracellular space.

Animals↗
PubMed1982

The structure of the cytoplasmic matrix preserved by freeze-drying and freeze-substitution.

This paper reports a study of the cytoplasmic matrix in whole cultured cells examined by high voltage electron microscopy. In order to acquaint ourselves with the influence of preparation procedures on the morphology depicted, PtK2 cells were prepared for examination by first freezing in propane at -185 degrees C and then drying while maintained at -95 degrees C. Other cells of the same type were first fixed with glutaraldehyde and OsO4 and then frozen dried. Others were prepared by freeze-substitution and eventually dried by the critical-point method from CO2. And finally, some were preserved by conventional techniques of glutaraldehyde and OsO4 followed by dehydration in alcohol and critical-point drying. The observations are presented in stereo images. The morphologies after these various procedures are quite similar. All show the characteristic three-dimensional lattice or meshwork of slender filaments called microtrabeculae. In cells rapidly frozen and then dried from the frozen state, there is less evidence of shrinkage and probable change in the trabecular structure than in cells first fixed with glutaraldehyde and then frozen dried. The differences we relate to the demonstrable failure of glutaraldehyde to penetrate the cell quickly and fix instantly any component that is in active motion. Other differences that can be observed between all four types of preparation are not recognizably striking and are thought to reflect as much as anything morphological diversity in the original cells. In some instances, the amorphous ice of the initial freezing at -185 degrees C was allowed to crystallize at -80 degrees C. The small crystals that form push aside the microtrabeculae and leave obvious imprints on the structure. The essential message from these experiments is that the cytomatrix is structured.

Animals↗
PubMed1981

Stabilization and the cytoplasmic ground substance in detergent-opened cells and a structural and biochemical analysis of its composition.

Treatment of epithelial BSC-1 cells with low concentrations of the detergent Brij 58 results in partial or complete removal of the plasmalemma and partial extraction of internal membrane-bound organelles without causing massive release of "cytosolic" proteins from the cytoplasmic ground substance. Stereoscopic high-voltage electron microscopy of such extracted and fixed cells demonstrates a system of slender (4-20 nm) strands in a three-dimensional "microtrabecular" arrangement similar to that observed in unextracted whole-mount preparations. Extraction of Brij-extracted cells with Triton X-100 dissolves many of the microtrabecular strands, leaving, as a more stable structure, a characteristic cytoskeletal network composed of various filaments and microtubules. Two-dimensional polyacrylamide gel electrophoresis of 35S-labeled polypeptides performed concurrently with the morphological studies demonstrates that Triton extraction of Brij-extracted cells releases a large number of polypeptides. This release parallels the loss of structural components observed by electron microscopy. Labeling of Brij-extracted cells with heavy meromyosin subfragment 1 decorates actin filaments with characteristic arrowhead complexes which are readily visualized only after subsequent Triton extraction. These observations support the concept that many cytoplasmic proteins are structure-bound and, in addition to the components comprising the cytoskeleton, are structure-forming. We conclude that a metastable association of various proteins of the cytoplasmic ground substance exists whose morphological integrity is maintained, at lest temporarily, after removal of the plasmalemma in solutions containing Brij 58.

Animals↗
PubMed1981

Localization and organization of actin in melanophores.

Melanophores of the angelfish, Pterophyllum scalare, were studied in an attempt to demonstrate the existence of actin in these cells although microfilaments had previously not been found. By use of a variety of procedures, including immunofluorescence microscopy of intact and detergent-extracted cells, transmission electron microscopy, high voltage electron microscopy of whole-mount preparations, and labeling with heavy meromyosin-subfragment 1, the presence of a loose cortical mesh of actin filaments is demonstrated. In addition, a more parallel array of filaments is detected in microspike- and microvillus-like surface projections. There seem to be no changes in the arrangement of these filaments as a function of the state of pigment distribution. No actin filaments could be found in association with pigment granules or microtubules in more central cell portions. For reasons presently unknown, the preservation of the cortical filament network in lysed cell preparations depends strongly on the presence of an intact microtubular system. The involvement of this subplasmalemmal actin filament network in pigment granule transport remains unclear.

Actins↗
PubMed1980

Observations on the innervated face of the electrocyte of the main organ of the electric eel (Electrophorus electricus L.).

The innervated face of electrocytes in the main electric organ of Electrophorus electricus L. was examined by light microscopy, both conventional and with Nomarski contrast, and by transmission and scanning electron microscopy. Acetylcholinesterase cytochemistry was used in the demonstration of the greater density of synapses over the caudal papillae. The various techniques contributed to a better understanding of the distribution and form of papillae and synapses at the posterior face of the electrocyte. Caudal papillae are longer and thinner than those at the rostral face, but it was not possible to recognize a different type sometimes referred to in the literature as small papillae. The contact of nerve endings with the electrocyte seems to be made predominantly on the terminal half of caudal papillae, however a smaller number occur elsewhere on the posterior face. Synaptic terminals frequently appear as round profiles, but may be also elongated, with or without bulges, usually occupying a depression, and separated from the post-synaptic membrane by a space of 60-100 nm, where an expansion may be found.

Acetylcholinesterase↗
PubMed1980

The control of pigment migration in isolated erythrophores of Holocentrus ascensionis (Osbeck). I. Energy requirements.

Erythrophores isolated from the scales of the marine teleost, Holocentrus ascensionis (Osbeck), are capable of rapidly aggregating or dispersing numberous red pigment granules within their cytoplasm by translocating them along radial paths delineated by bundles of radially oriented microtubules. Pigment translocation is accompanied by transformations in the morphology of the cytoplasmic matrix, or microtrabecular lattice (MTL), in which the pigment granules are suspended. It appears that the MTL as a whole contracts toward the cell center during aggregation, carrying the pigment granules inward along with it, and is restructured during dispersion, using the radial microtubules as guides. We examined the energy requirements of pigment migration and the accompanying MTL transformations. Cellular ATP was depleted using the specific metabolic inhibitors 2,4 dinitrophenol, NaCN and oligomycin. All three of these drugs, which inhibit oxidative phosphorylation by different mechanisms, prevent both pigment dispersion and MTL transformation to dispersed morphology, while aggregation is unaffected. Inhibitor-treated cells recover normal pigment movements and MTL morphology when inhibitor is washed out of the cells with fresh medium. Potential energy apparently is stored in the MTL by some ATP-dependent process during dispersion and is converted to kinetic energy during aggregation. The results of this study strengthen the hypothesis that the MTL, working in concert with the radial microtubules, is the vehicle for pigment translocation in the erythrophore system.

Adenosine Triphosphate↗
PubMed1980

Visualization of microtubules of cells in situ by indirect immunofluorescence.

Microtubule staining patterns can be visualized within cells in situ on the surface of fish scales from the squirrel fish, Holocentrus ascensionis, and the common goldfish, Carassius auratus, after incubation with antibodies to sea urchin tubulin and fluorescein-labeled goat antibodies to rabbit immunoglobulin G. Chromatophores in situ from both species reveal a radial microtubule framework that orients the alignment of pigment granules. Innervating fibers of erythrophores on the H. ascensionis scale can also be observed. In situ, pseudo-epithelial cells called scleroblasts show microtubule patterns with a remarkable degree of similarity within a selected region. Over 90% of the cells have a microtubule framework that is nearly superimposable from cell to adjacent cell. The microtubules in scleroblasts are few and form a simple radial framework with a localized microtubule organizing center (MTOC). Microtubules in scleroblasts in vitro emanate from localized MTOCs but are much less radially organized than in situ. Scleroblasts in situ on the scale of C. auratus show microtubules that curve abruptly into coalignment with phase striations on the fibrillary plate. The phase striations arise from the orthogonal plies of collagen in intimate association with the scleroblasts. The role of microtubules in scleroblasts may thus be to provide orientation for collagen fibrillogenesis, analogous to their role in orientation of cellulose fibers in plants. That cells in situ exhibit highly related and coordinated microtubule staining patterns reaffirms that the cytoskeleton plays an important role in the organization of differentiated tissues.

Animals↗
PubMed1980

Microtrabecular structure of the axoplasmic matrix: visualization of cross-linking structures and their distribution.

Axoplasmic transport is a dramatic example of cytoplasmic motility. Constituents of axoplasm migrate as far as 400 mm/d or at approximately 5 micron/s. Thin-section studies have identified the major morphological elements within the axoplasm as being microtubules, neurofilaments (100-A filaments), an interconnected and elongated varicose component of smooth endoplasmic reticulum (SER), more dilated and vesicular organelles resembling portions of SER, multivesicular bodies, mitochondria, and, finally, a matrix of ground substance in which the tubules, filaments, and vesicles are suspended. In the ordinary thin-section image, the ground substance is comprised of wispy fragments which, in not being noticeably tied together, do not give the impression of representing more than a condensation of what might be a homogeneous solution of proteins. With the high-voltage microscope on thick (0.5-micron) sections, we have noticed, however, that the so-called wispy fragments are part of a three-dimensional lattice. We contend that this lattice is not an artifact of aldehyde fixation, and our contention is supported by its visability after rapid-freezing and freeze-substitution. This lattice or microtrabecular matrix of axoplasm was found to consist of an organized system of cross-bridges between microtubules, neurofilaments, cisternae of the SER, and the plasma membrane. We propose that formation and deformation of this system are involved in rapid axonal transport. To facilitate electron microscope visualization of the trabecular connections between elements of axoplasm, the following three techniques were used: first, the addition of tannic acid to the primary fixative, OsO4 postfixation, then en bloc staining in uranyl acetate for conventional transmission electron microscope (TEM); second, embedding tissue in polyethylene glycol for thin sectioning, dissolving out the embedding medium from the sections and blocks, critical-point-drying (J. J. Wolosewick, 1980, J. Cell Biol., 86:675-681.), and then observing the matrix-free sections with TEM or the blocks with a scanning electron microscope; and third, rapid freezing of fixed tissue followed by freeze-etching and rotary-shadowing with replicas observed by TEM. All of these procedures yielded images of cross-linking elements between neurofilaments and organelles of the axoplasm. These improvements in visualization should enable us to examine the distribution of trabecular links on motile axonal organelles.

Animals↗
PubMed1979

Microtrabecular lattice of the cytoplasmic ground substance. Artifact or reality.

The cytoplasmic ground substance of cultured cells prepared for high voltage transmission electron microscopy (glutaraldehyde/osmium fixed, alcohol or acetone dehydrated, critical-point dried) consists of slender (3-6 nm Diam) strands--the microtrabeculae (55)--that form an irregular three-dimensional lattice (the microtrabecular lattice). The microtrabeculae interconnect the membranous and nonmembranous organelles and are confluent with the cortices of the cytoplast. The lattice is found in all portions of the cytoplast of all cultured cells examined. The possibility that the lattice structure is an artifact of specimen preparation has been tested by (a) subjecting whole cultured cells (WI-38, NRK, chick embryo fibroblasts) to various chemical (aldehydes, osmium tetroxide) and nonchemical (freezing) fixation schedules, (b) examination of model systems (erythrocytes, protein solutions), (c) substantiating the relaibility of critical-point drying, and (d) comparing images of whole cells with conventionally prepared (plastic-embedded) cells. The lattice structure is preserved by chemical and nonchemical fixation, though alterations in ultrastructure can occur especially after prolonged exposure to osmium tetroxide. The critical-point method for drying specimens appears to be reliable as is the freeze-drying method. The discrepancies between images of plastic-embedded and sectioned cells, and images of whole, critical-point dried cells appear to be related, in part, to the electron-scattering properties of the embedding resin. The described observations indicate that the microtrabecular lattice seen in electron micrographs closely represents the nonrandom structure of the cytoplasmic ground substance of living cultured cells.

Animals↗
PubMed1977

Observation on the morphological heterogeneity of WI-38 cells.

WI-38 cells of intermediate and late-passage cultures were examined by light microscopy, scanning, conventional transmission and high voltage electron microscopy for evidence of heterogeneity among the cells within a single culture. Two morphologically distinct cell sizes and shapes were noted in all passages, (1) a typical fibroblastic type, and (2) a much larger, non-fusiform type. The larger cells generally had a nucleus that was positioned to one side of the bulk of the cytoplasm. The smaller was consistently fusiform with a centrally placed nucleus. The surfaces of intermediate-passage cells were uniform in showing small microvilli and corticall pits but were otherwise smooth and with out distinctive featues. The late-passage cells, on the other band, were consistent in showing numerous blebs and marginal ruffing. The internal structure of these cells in all passages studied was complicated by many age-related changes. The observations indicate that there is in WI-38 cells a high degree of intraculture heterogeneity. An awareness of this is important in studies which characterize the biochemical properties of this cell strain.

Cell Line↗
PubMed1977

Transformations in the structure of the cytoplasmic ground substance in erythrophores during pigment aggregation and dispersion. I. A study using whole-cell preparations in stereo high voltage electron microscopy.

Pigment migration in cultured erythrophores of the squirrel fish Holocentrus ascensionis, after manipulation with K+, epinephrine, 3',5'-dibutyryl cyclic adenosine monophosphate, theophylline, and caffeine, is essentially identical to that observed in this chromatophore in situ. For such observations, the erythrophores are dissociated from the scales with hyaluronidase and collagenase, and allowed to spread on an amorphous collagen substrate, where they resemble the discoid erythrophore in situ. In this state, they are readily fixed by glutaraldehyde and osmium tetroxide, and are then critical-point dried for whole-cell viewing in the high voltage electron microscope. The organization and fine structure of the erythrophore cytoplast was stereoscopically examined after fixation of the pigment granules in four experimental states: pigment dispersed, pigment aggregated, pigment aggregating, and pigment dispersing. In the dispersed cell, granules are contained in an extensive three-dimensional lattice composed of radially oriented microtubules and a network of fine filaments 3-6 nm in diameter (microtrabeculae), whereas in the aggregated cell, the microtrabecular system is absent, and the majority of the microtubules appear displaced into the cortices on the cytoplasmic surface of the plasma membrane. In cells fixed while aggregating, few microtrabeculae are observed, although formless thickenings are observed in the cortices, on granules, and between clumped granules. In dispersing cells, the microtrabecular system is reformed from material stored in the cortices and with the granules in the centrosphere. These observations suggest that the granules are suspended in a dynamic microtrabecular system that withdraws during pigment aggregation and is restructured during pigment dispersion. The microtubules guide linear granule motion not by defining physical channels, but by a recognizable affinity of microtubules, microtrabeculae, and granules for one another.

Animals↗